Bonded-phase silicas commonly used in chromatographic applications have been described in a variety of publications and are well-known in the art. The preparation of these stationary phases has been described using monofunctional, bifunctional and trifunctional silanes. The bonded silicas described however, did not use sterically-protecting groups, and thus have been found to be relatively unstable over a wide variety of operating conditions.
In general a reverse phase chromatography column containing a bonded silica as the stationary phase is stable over a relatively narrow pH range. Frequently the pH must be maintained at a prescribed pH, or the column undergoes irreversible damage losing its efficiency and characteristics, such as the ability to produce narrow peaks, desirable retention volumes or resolve components of a mixture. This damage can occur even if the mistaken use outside the narrow pH range defined for the column is only for a short period of time. In many cases, these stationary phases are produced by reacting an alkyldimethylchlorosilane, alkyldimethylalkoxysilane, or alkyltrialkyloxysilane with a silica in the presence of a base such as pyridine to form a siloxane bond to the surface.
In U.S. Pat. No. 4,705,725, a stable support structure comprising a substrate to which is attached a monofunctional silane containing two sterically-protecting groups R and an additional functional group R' is disclosed. The compounds disclosed in the patent are represented by the following Formula (A): ##STR2## where R=isopropyl, t-butyl, sec-butyl, sec-pentyl, isopentyl or cyclohexyl; R'=alkane, substituted alkane, alkene, substituted alkene, aryl or substituted aryl and A=a surface group of a substrate to which the silane is covalently attached. The sterically-protecting R groups were found to impart surprising stability to these support structures. These silanes are produced by the hydrosilation of an alkene by chloro-di-(2-propyl)silane which uses an expensive catalyst and generally occurs in a relatively low quantitative yield requiring separation of the product from the starting materials.
In the specification of the '725 Patent, it is taught that the silicon atoms within the silane of this invention must be monofunctional (col 5-6, lines 66-68 and 1-16, respectively). It is taught that multifunctional reagents will tend to polymerize during reaction or use, resulting in irreproducible surfaces with different chemical and physical characteristics. It is also taught that multifunctional silanes only partially react with a surface, leaving a reactive group that can ultimately undergo unwanted reactions with materials being handled or hydrolyze to acidic silanols that can deleteriously interact with materials of interest.
It has now been unexpectedly found that bifunctional silanes containing two-sterically protective groups can be used to produce reproducible and stable covalently-bonded silanes. The synthesis of these silanes does not require a hydrosilation step and permits the preparation of a wide variety of functional silanes displaying stability over a wide range of pHs.